U.S. patent application number 11/890321 was filed with the patent office on 2008-01-24 for hemostatic compositions and methods for controlling bleeding.
This patent application is currently assigned to Crosslink-D, Inc.. Invention is credited to Kent C. Cochrum, Susan Jemtrud.
Application Number | 20080020026 11/890321 |
Document ID | / |
Family ID | 26993389 |
Filed Date | 2008-01-24 |
United States Patent
Application |
20080020026 |
Kind Code |
A1 |
Cochrum; Kent C. ; et
al. |
January 24, 2008 |
Hemostatic compositions and methods for controlling bleeding
Abstract
The invention provides hemostatic compositions useful to promote
hemostasis at active bleeding wound sites. The hemostatic
compositions typically include an article containing cellulose,
e.g., cotton gauze, and a polysaccharide covalently linked to the
cellulose, or a polysaccharide ionically cross-linked and in
association with the article. Methods of making and using the
hemostatic compositions are also provided.
Inventors: |
Cochrum; Kent C.; (West
Sacramento, CA) ; Jemtrud; Susan; (Auburn,
CA) |
Correspondence
Address: |
FISH & RICHARDSON P.C.
PO BOX 1022
MINNEAPOLIS
MN
55440-1022
US
|
Assignee: |
Crosslink-D, Inc.
West Sacramento
CA
|
Family ID: |
26993389 |
Appl. No.: |
11/890321 |
Filed: |
August 6, 2007 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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11357770 |
Feb 17, 2006 |
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11890321 |
Aug 6, 2007 |
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10334864 |
Dec 31, 2002 |
7101862 |
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11357770 |
Feb 17, 2006 |
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60343247 |
Dec 31, 2001 |
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60354917 |
Feb 11, 2002 |
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Current U.S.
Class: |
424/446 ;
424/488; 424/489 |
Current CPC
Class: |
A61L 15/225 20130101;
A61L 15/44 20130101; A61L 15/28 20130101; A61L 15/225 20130101;
A61L 15/28 20130101; A61L 2400/04 20130101; A61L 15/28 20130101;
C08L 1/26 20130101; A61L 15/28 20130101; C08L 5/02 20130101; C08L
5/04 20130101; A61P 7/04 20180101; C08L 5/04 20130101; A61L
2300/232 20130101; A61L 33/08 20130101; A61L 2300/418 20130101 |
Class at
Publication: |
424/446 ;
424/488; 424/489 |
International
Class: |
A61K 9/70 20060101
A61K009/70; A61K 9/14 20060101 A61K009/14; A61P 7/04 20060101
A61P007/04; A61K 9/16 20060101 A61K009/16 |
Claims
1. A method for controlling bleeding at an active bleeding wound
site of an mammal, said method comprising applying a hemostatic
composition to said active bleeding wound site, said hemostatic
composition comprising an article comprising cellulose and a
polysaccharide covalently linked to said cellulose.
2. The method of claim 1, wherein said polysaccharide is selected
from the group consisting of cross-linked dextran, cross-linked
starch, and cross-linked alginate.
3. The method of claim 2, wherein said polysaccharide is
cross-linked dextran.
4. The method of claim 2, wherein the molecular weight of said
cross-linked dextran is from about 10,000 to about 2,000,000.
5. The method of claim 4, wherein the molecular weight of said
cross-linked dextran ranges from about 20,000 to about 100,000.
6. The method of claim 1, wherein said covalently linked
polysaccharide has a molecular weight exclusion limit greater than
about 30,000 Daltons.
7. The method of claim 1, wherein said article comprising cellulose
is selected from the group consisting of a bandage, suture,
dressing, gauze, gel, foam, web, film, tape, or patch.
8. The method of claim 1, wherein said article comprising cellulose
comprises cotton.
9. The method of claim 8, wherein said article comprising cellulose
is cotton gauze.
10. The method of claim 1, wherein prior to said application of
said hemostatic composition, said active bleeding wound site bleeds
at a rate from about 0.5 ml/min to about 1000 ml/min.
11. The method of claim 1, wherein after application of said
hemostatic composition, said active bleeding wound site bleeds at a
rate of less than 0.03 ml/min.
12. The method of claim 11, wherein said rate of less than 0.03
ml/min is achieved from about 2 to about 20 minutes after
application of said hemostatic composition.
13. The method of claim 1, wherein said hemostatic composition
further comprises a second polysaccharide covalently linked to said
cellulose.
14. The method of claim 5, wherein said hemostatic composition
further comprises a second polysaccharide, wherein said second
polysaccharide is cross-linked dextran having a molecular weight
from about 800,000 to about 2M, wherein said second polysaccharide
is covalently linked to said cellulose.
15. A method for controlling bleeding at an active bleeding wound
site of an animal, said method comprising applying a hemostatic
composition to said active bleeding wound site, said hemostatic
composition comprising an article comprising cellulose in
association with ionically cross-linked alginate.
16. The method according to claim 15, wherein said alginate is
ionically cross-linked with a cation selected from the group
consisting of: Mg2+; Ni2+; Ca2+; Sr2+; Ba2+; Zn2+; Cd2+; Cu2+;
Pb2+; Fe3+; and Al3+.
17. The method according to claim 15, further comprising a second
polysaccharide covalently linked to said cellulose, wherein said
second polysaccharide is cross-linked dextran.
18. The method according to claim 17, wherein said alginate is
further covalently linked to said cross-linked dextran.
19. The method of claim 1, wherein said mammal is a human.
20. The method of claim 3, wherein said cross-linked dextran is in
the form of beads.
Description
CROSS REFERENCED TO RELATED APPLICATION
[0001] This application is a continuation of U.S. patent
application Ser. No. 11/357,770 filed Feb. 17, 2006, which is a
divisional of U.S. patent application Ser. No. 10/334,864 filed
Dec. 31, 2002, which claims priority from U.S. Provisional Patent
Application No. 60/343,247, filed Dec. 31, 2001, and from U.S.
Provisional Patent Application No. 60/354,917, filed Feb. 11, 2002,
all of which are incorporated herein by reference in their
entirety.
TECHNICAL FIELD
[0002] This invention relates to hemostatic compositions and
methods employing the same, and more particularly to hemostatic
compositions useful for controlling bleeding at active bleeding
wound sites.
BACKGROUND
[0003] Wounds are generally classified as acute or chronic in
accordance with their healing tendencies. Acute wounds, typically
those received as a result of surgery or trauma, usually heal
uneventfully within an expected time frame. Acute wounds include
wounds such as active bleeding wound sites, e.g., wounds that have
detectable, unclotted blood. The rapid control of topical bleeding
at active bleeding wound sites is of critical importance in wound
management, especially for the management of trauma, e.g., as a
result of military exercises or surgery.
[0004] A conventional method of controlling bleeding at active
bleeding wound sites, such as an external hemorrhage or a surgical
wound, advocates the use of cotton gauze pads capable of absorbing
250 ml of blood. Cotton pads are considered passive, however,
because of their inability to initiate or accelerate blood
clotting. Other formulations have been reported to promote
hemostasis and are described in U.S. Pat. Nos. 6,454,787;
6,060,461; 5,196,190; 5,667,501; 4,793,336; 5,679,372; 5,098,417;
and 4,405,324. A hemostatic composition capable of accelerating the
coagulation cascade to form a thrombus would be useful.
SUMMARY
[0005] Accordingly, the invention relates to hemostatic
compositions and methods for making and using the same in order to
promote hemostasis at active bleeding wound sites. The present
compositions typically include an article which contains cellulose,
e.g., cotton gauze, and a polysaccharide covalently linked to the
cellulose. In other embodiments, a polysaccharide is ionically
cross-linked and in association with an article comprising
cellulose. Hemostatic compositions can include additional
polysaccharides covalently linked to either or both of the
cellulose and the first polysaccharide or physically trapped by a
network formed by the covalent linking or ionic cross-linking of
the first polysaccharide.
[0006] In one aspect of the invention, a method for controlling
bleeding at an active bleeding wound site of an animal is provided.
The animal can be a mammal. For example, the animal can be a human,
horse, bird, dog, cat, sheep, cow, or monkey. The method includes
applying a hemostatic composition to the active bleeding wound
site. The hemostatic composition includes an article which contains
cellulose and a polysaccharide, such as dextran, starch, or
alginate, covalently linked to the cellulose. If dextran is used,
it may be in the form of a bead, e.g., covalently cross-linked
dextran beads. The molecular weight of the dextran can range from
about 10,000 to about 2,000,000 Daltons, or from about 20,000 to
about 100,000 Daltons. When a polysaccharide is linked to the
cellulose, it can have a molecular weight exclusion limit of
greater than about 30,000 Daltons.
[0007] Articles which contain cellulose can be barriers,
structures, or devices useful in surgery, diagnostic procedures, or
wound treatment. For example, an article containing cellulose can
be a bandage, suture, dressing, gauze, gel, foam, web, film, tape,
or patch. An article containing cellulose can include a cotton
material, e.g., cotton gauze. The article can also optionally
include adhesives or polymeric laminating materials.
[0008] Hemostatic compositions of the present invention are useful
for accelerating blood clotting at an active bleeding wound site.
Prior to the application of a hemostatic composition, an active
bleeding wound site may be characterized in that it bleeds at a
rate of from about 0.5 ml/min to about 1000 ml/min. After
application of a hemostatic composition, the active bleeding wound
site may bleed at a rate of less than 0.03 ml/min. For example, the
rate of less than 0.03 ml/min. may be achieved in from about 2 to
about 20 minutes, and in certain embodiments in less than about 5
minutes.
[0009] A hemostatic composition can comprise a second
polysaccharide covalently linked to the cellulose and, optionally,
to the first polysaccharide. The second polysaccharide may have a
different molecular weight than the first polysaccharide. For
example, the second polysaccharide may be dextran having a
molecular weight from about 800,000 to about 2M. Alternatively, the
second polysaccharide may be physically trapped by the covalent
linking of the first polysaccharide to the cellulose.
[0010] In other embodiments, hemostatic compositions of the present
invention can include an article comprising cellulose in
association with a polysaccharide ionically linked to itself
(cross-linked). For example, the article comprising cellulose may
be coated with, immersed in, or soaked in the polysaccharide, which
is subsequently ionically cross-linked. The polysaccharide may be
further covalently linked to the cellulose of the article. In
addition, in certain embodiments, the polysaccharide may be
physically trapped in fibers of the article comprising
cellulose.
[0011] One example of a polysaccharide that can be ionically
cross-linked is alginate. Alginate can be ionically cross-linked to
itself with metal cations, including Mg2+; Ni2+; Ca2+; Sr2+; Ba2+;
Zn2+; Cd2+; Cu2+; Pb2+; Fe3+; and Al3+. In some embodiments, the
cation is Ca2+. A second polysaccharide, such as dextran, can also
be physically trapped, e.g., by the network formed by the ionic
cross-linking of the first polysaccharide. Dextran can be in the
form of cross-linked beads, e.g., dextran that has been previously
cross-linked to itself. Dextran can be covalently linked to the
bandage, e.g. by linking dextran to the cellulose with
epichlorohydrin.
[0012] In another aspect, a hemostatic composition can include
dextran-alginate spheres, such as ionically linked dextran-alginate
spheres, or covalently linked dextran-alginate spheres, or both
ionically and covalently linked dextran-alginate spheres.
[0013] In another aspect of the invention, hemostatic compositions
are provided that include additional agents, such as analgesics,
steroids, antihistamines, anesthetics, bactericides, disinfectants,
fungicides, vasoconstrictors, hemostatics, chemotherapeutic drugs,
antibiotics, keratolytics, cauterizing agents, antiviral drugs,
epidermal growth factor, fibroblast growth factors, transforming
growth factors, glycoproteins, collagen, fibrinogen, fibrin,
humectants, preservatives, lymphokines, cytokines, odor controlling
materials, vitamins, and clotting factors.
[0014] The invention also provides methods for making hemostatic
compositions. Hemostatic compositions of the present invention can
be made by incubating a linking agent with a polysaccharide and an
article comprising cellulose to form a hemostatic composition
having the polysaccharide covalently linked to the cellulose.
[0015] The linking agent may be any linking agent useful for
linking available hydroxyl groups on cellulose with available
hydroxyl groups on a polysaccharide. Examples include
epichlorohydrin, dichlorohydrin, diepoxyburan, disepoxypropyl
ether, or ethylene-glyco-bis-epoxypropylether. The incubation step
may occur in an aqueous alkaline solution. The temperature of the
incubation step can range from about 40.degree. C. to about
70.degree. C. In certain embodiments, the temperature is about
50.degree. C. The incubation step can occur for about 1 to about 24
hours. In addition, the incubation step can be in the presence of a
stabilizing solution, e.g., a solution designed to prevent or limit
evaporation of water. The stabilizing solution can include
cellulose acetate butyrate. The covalently linked polysaccharide
may have a molecular weight exclusion limit of greater than 30,000
Daltons.
[0016] In certain embodiments of the method, the polysaccharide is
dextran. The dextran can be in the form of covalently cross-linked
beads. The molecular weight of the dextran can range from about
10,000 to about 2M, or from about 20,000 to about 100,000 Daltons.
The incubation step may be occur in an aqueous alkaline solution
having about 12% to about 75% dextran.
[0017] In another aspect, the invention provides a method of making
a composition including incubating a polysaccharide and a cation
with an article containing cellulose in order to form a hemostatic
composition having the article containing cellulose in association
with an ionically cross-linked polysaccharide. The polysaccharide
may be further covalently linked to the cellulose. The cation may
be, for example, Ca2+. The Ca2+ may be in the form of, or derived
from, Ca2+-loaded cross-linked dextran beads. The polysaccharide
may be sodium alginate or a derivative of alginic acid, including
salts of alginic acid.
[0018] In certain embodiments, the incubation step includes a
second polysaccharide; the second polysaccharide may become
physically trapped in the three dimensional network formed by the
ionic cross-linking of the first polysaccharide. The second
polysaccharide may be dextran, e.g., dextran in the form of
cross-linked beads. The second polysaccharide may be further
covalently linked to the cellulose, e.g., through a linking agent
such as epichlorohydrin.
[0019] In another aspect, the invention provides a method for
manufacturing a composition, where the method includes the step of
mixing an aqueous phase alkaline polysaccharide solution with an
organic phase stabilizing agent solution to form a mixture having
polysaccharide spheres; incubating a cross-linking agent with the
mixture to cross-link the polysaccharide spheres; isolating the
cross-linked polysaccharide spheres; and coating an article
comprising a sodium alginate solution with the cross-linked
polysaccharide spheres. The method can include removing the organic
phase stabilizing agent from the mixture, e.g., prior to isolating
the cross-linked polysaccharide spheres. The method can also
include exposing the cross-linked polysaccharide spheres to a
solution comprising Ca2+ ions, e.g., washing the cross-linked
polysaccharide spheres in a Ca2+ solution. The polysaccharide may
be dextran, and the organic phase stabilizing agent solution may
include cellulose acetate butyrate.
[0020] In certain embodiments of the method, cross-linked
polysaccharide spheres are between about 30 to about 500 .mu.m in
size. The mixing and incubating steps may occur at a temperature of
from about 40.degree. C. to about 70.degree. C. The coating step
can include spraying the article with the cross-linked
polysaccharide spheres. The invention also relates to hemostatic
compositions manufactured according to the above method.
[0021] In a further aspect, another method for manufacturing a
composition is provided. The method includes the steps of providing
an aqueous phase alkaline solution having dextran and sodium
alginate therein; preparing dextran-alginate spheres from the
aqueous phase alkaline solution; and incubating the
dextran-alginate spheres with a linking agent to link said
dextran-alginate spheres. Dextran-alginate spheres can be prepared
by any method conventional in the art, including the use of a
mechanical droplet generator. A linking agent may covalently or
ionically link or cross-link the dextran-alginate spheres.
Accordingly, a linking agent may be epichlorohydrin or a
Ca2+-containing salt such as calcium chloride. The dextran-alginate
spheres may be linked with a Ca2+ linking agent, and then linked
with an epichlorohydrin linking agent, or the linking can be
performed in the reverse order, or simultaneously. The method can
further including coating, e.g., spraying, an article, such as an
article comprising sodium alginate, with the linked
dextran-alginate spheres. The invention also includes hemostatic
compositions manufactured according to the method.
[0022] Unless otherwise defined, all technical and scientific terms
used herein have the same meaning as commonly understood by one of
ordinary skill in the art to which this invention belongs. Although
methods and materials similar or equivalent to those described
herein can be used to practice the invention, suitable methods and
materials are described below. All publications, patents, patent
applications, and other references mentioned herein are
incorporated by reference in their entirety. In case of conflict,
the present specification, including definitions, will control. In
addition, the materials, methods, and examples are illustrative
only and not meant to be limiting.
[0023] The details of one or more embodiments of the invention are
set forth in the accompanying drawings and the description below.
Other features, objects, and advantages of the invention will be
apparent from the description and drawings, and from the
claims.
DESCRIPTION OF DRAWINGS
[0024] Like reference symbols in the various drawings indicate like
elements.
DETAILED DESCRIPTION
[0025] As used herein, the terms "linking" or "linked" are meant to
indicate either a covalent or ionic link, either direct or mediated
by a chemical moiety or an ion, between two chemically distinct
entities, e.g., dextran linked to cellulose. The term "cross-link"
is meant to indicate a covalent or ionic link, either direct or
mediated by a chemical moiety or ion, between two chemically
similar moieties, e.g., dextran cross-linked to itself, alginate
cross-linked to itself. The chemically similar moieties do not have
to be identical. For example, dextran having a particular average
molecular weight range includes dextran molecules of a variety of
molecular weights, and thus the dextran molecules are not identical
but chemically similar. When dextran molecules having an average
molecular weight range are linked, e.g., covalently linked with
epichlorohydrin, they are said to be "cross-linked."
[0026] The terms "spheres," "particles," or "beads," when used in
the context of the present invention, are not meant to imply
different sizes, but are meant to be interchangeable terms
describing an embodiment of a composition.
[0027] The term "active bleeding wound site" means, at a minimum,
that unclotted blood is present in the wound, e.g., extravascular
blood, particularly where the surface of a tissue has been broken
or an artery, vein, or capillary system has been compromised. The
rate of blood flow from an active bleeding wound site can vary,
depending upon the nature of the wound. In some cases, an active
bleeding wound site will exhibit blood flow at a rate from about at
a rate of from about 0.5 ml/min to about 1000 ml/min. Some active
bleeding wound sites may exhibit higher rates of blood flow, e.g.,
punctures of major arteries such as the aorta. After application of
the hemostatic composition, the active bleeding wound site may
bleed at a rate of less than 0.03 ml/min. For example, the rate of
less than 0.03 ml/min. may be achieved in from about 2 to about 20
minutes, and in certain embodiments in less than about 5
minutes.
[0028] Hemostatic Compositions
[0029] The invention relates to hemostatic compositions used to
promote hemostasis at active bleeding wound sites. While not being
bound by any theory, it is believed that the hemostatic
compositions of the present invention control bleeding by
initiating and accelerating blood clotting. The hemostatic
compositions of the present invention activate platelets and
concentrate high molecular weight components of the coagulation
cascade (e.g., clotting factors) by excluding high molecular weight
components of the cascade, while absorbing the lower molecular
weight components in blood. Accordingly, coagulation cascade
components having a molecular weight higher than about 30,000
Daltons are excluded, including fibrinogen (MW 340,000);
prothrombin (MW 70,000); thrombin (MW 34,000); Factor V (MW
330,000); Factor VII (MW 50,000); Factor VIII (MW 320,000); von
Willebrand factor (MW>850,000); Factor IX (MW 57,000); Factor X
(MW 59,000); Factor XI (MW 143,000); Factor XII (MW 76,000); Factor
XIII (MW 320,000); high MW kininogen (Fitzgerald Factor) (MW
120,000-200,000), and prekallikrein (Fletcher Factor) (MW
85,000-100,000). In addition, laboratory experiments indicate that
platelets aggregate around the hemostatic compositions of the
present invention when exposed to blood. The net result is that
concentrated clotting factors (coagulation cascade components) and
activated platelets activate the conversion of prothrombin to
thrombin in the presence of Ca2+, which subsequently catalyzes the
conversion of fibrinogen to insoluble fibrin multimers, e.g., a
fibrin clot. Additional information on the clotting cascade and
hemostatic compositions containing fibrin can be found in U.S. Pat.
No. 5,773,033.
[0030] Hemostatic compositions typically include an article
comprising cellulose, e.g., cotton gauze, and a polysaccharide
covalently linked to the cellulose. In other embodiments,
hemostatic compositions include an article comprising cellulose in
association with a polysaccharide that is ionically cross-linked.
The polysaccharide can be further covalently linked to the
cellulose. Hemostatic compositions can include additional
polysaccharides covalently linked to either or both of the
cellulose and the first polysaccharide. Other embodiments of
hemostatic compositions include linked and cross-linked
polysaccharide spheres, optionally loaded with a cation, e.g.,
Ca2+.
[0031] It should be noted that certain hemostatic compositions
comprise both a macroscopic structure (e.g., an article) and a
microscopic structure (e.g., networks of polysaccharide
cross-linkages or networks of polysaccharide covalent linkages to
cellulose). Some hemostatic compositions therefore form three
dimensional networks of a polysaccharide, either as ionically
linked chains or covalently bound to the cellulose of the article.
Accordingly, in some embodiments, a second polysaccharide may be
physically trapped by the network formed by the first
polysaccharide.
[0032] Accordingly, in one aspect, a hemostatic composition
includes an article containing cellulose and a polysaccharide, such
as dextran, starch, or alginate, covalently linked to the
cellulose. The article may include natural or synthetic celluloses
(e.g., cellulose acetate, cellulose butyrate, cellulose
propionate). The polysaccharide chosen should be safe for in vivo
use, e.g., non-allergenic, non-toxic, and preferably
non-metabolized. Polysaccharides for clinical use are known in the
art and available from a variety of sources. See, e.g., U.S. Pat.
No. 6,303,585.
[0033] As used herein, covalent linkages encompass bonds from any
of the available chemical moieties of the polysaccharide to any of
the available chemical moieties of the cellulose. For example, if
the polysaccharide dextran is used, hydroxyl moieties on dextran
can be covalently linked to hydroxyl moieties on cellulose through
the linking agent epichlorohydrin. In that case, a glyceryl bridge
linking dextran to cellulose is formed. For additional information,
see Flodin, P., and Ingelman, B., "Process for the Manufacture of
Hydrophilic High Molecular Weight Substances," British Patent No.
854,715; and Flodin, P. "Chapter 2: The Preparation of Dextran
Gels," Dextran Gels and Their Applications in Gel Filtration,
Pharmacia, Uppsala Sweden, 1962, pages 14-26.
[0034] The average molecular weight range of the polysaccharide can
vary, but typically ranges from about 10,000 to about 2M Daltons.
The molecular weight range chosen will affect the molecular weight
exclusion limit of the covalently linked polysaccharide, and thus
its ability to exclude the coagulation components and concentrate
them.
[0035] Dextran is a high molecular weight polysaccharide that is
water-soluble. It is not metabolized by humans, and is non-toxic
and tolerated well by most animals, including humans. The average
molecular weight of dextran used in the present invention can range
from about 10,000 to about 2,000,000 Daltons, or from about 20,000
to about 100,000 Daltons.
[0036] Dextran can be in the form of beads, e.g., covalently
cross-linked beads, before it is linked covalently to the
cellulose. Dextran beads can exhibit a range of sizes, e.g., from
about 30 to about 500 .mu.m. Dextran beads are commercially
available, e.g., as Sephadex.TM. (Pharmacia); see, for example UK
974,054. Alternatively, dextran beads or particles may be formed
during the preparation of the hemostatic composition, e.g., from
the covalent cross-linking of previously uncross-linked dextran
molecules.
[0037] In other embodiments, dextran may be in solution form, e.g.,
uncross-linked, before it is covalently linked to the cellulose.
Dextran may be covalently linked to the cellulose and covalently
cross-linked to itself, e.g., when exposed to a linking agent such
as epichlorohydrin. When dextran is in solution form (e.g.,
uncross-linked), the dextran molecules may coat all or a component
of the article, such as fibers of a cotton bandage, so that it
subsequently forms a three-dimensional microscopic linked network
or mesh when it is covalently linked to the cellulose and
covalently cross-linked to itself. Dextran beads linked to
cellulose or a cellulose-dextran mesh as described previously
contribute to the ability of a hemostatic composition to exclude
high molecular weight components of the coagulation cascade.
[0038] The average molecular weight of the polysaccharide, the
degree of linking of the polysaccharide to cellulose, and any
cross-linking of the polysaccharide (e.g., to itself) are factors
in the molecular weight exclusion limit of the polysaccharide in a
hemostatic composition and the water regain of a hemostatic
composition. Water regain is defined as the weight of water taken
up by 1 g of dry hemostatic composition and can be determined by
methods known in the art. For example, it is known that small
changes in dextran concentration or linking agent concentration
(e.g., epichlorohydrin) can result in dramatic changes in water
regain. Typically, at lower molecular weights of dextran, a higher
water regain results. See Flodin, P., "Chapter 2: The Preparation
of Dextran Gels," Dextran Gels and Their Applications in Gel
Filtration, Pharmacia, Uppsala Sweden, 1962, pages 14-26.
[0039] Similarly, the degree of hydration of the polysaccharide
also affects the molecular weight exclusion limit. As the degree of
hydration increases, the molecular weight exclusion limit of the
polysaccharide usually increases. Typically, when dextran is linked
to cellulose, the dextran will have a molecular weight exclusion
limit of greater than about 30,000 Daltons, thus effectively
excluding the components of the coagulation cascade and
concentrating them on the microscopic surface of the hemostatic
composition.
[0040] Articles which contain cellulose can be any barriers,
structures, or devices useful in surgery, diagnostic procedures, or
wound treatment. For example, an article containing cellulose can
be a bandage, suture, dressing, gauze, gel, foam, web, film, tape,
or patch. An article containing cellulose can include a cotton
material, e.g., cotton gauze. The article should allow the
polysaccharide linked to the cellulose to interact with the wound
site.
[0041] A hemostatic composition can comprise a second
polysaccharide covalently linked to cellulose. The second
polysaccharide may have a different molecular weight than the first
polysaccharide. For example, the second polysaccharide may be
dextran having a molecular weight from about 800,000 to about 2M.
The second polysaccharide may be covalently linked to the cellulose
at a time after the first polysaccharide, at the same time as the
first polysaccharide, or at a time before the first
polysaccharide.
[0042] In other embodiments, hemostatic compositions of the present
invention can include an article containing cellulose in
association with an ionically cross-linked polysaccharide. The
polysaccharide may be further covalently linked to the cellulose.
In this context, ionic linkages include ion-mediated bonds between
available chemical moieties on the polysaccharide. Typical chemical
moieties that can be mediated with an ion (e.g., a cation) include
hydroxyl moieties. For example, sodium alginate or alginic acid
salts can be ionically linked with metal cations, including Mg2+,
Ni2+, Ca2+, Sr2+, Ba2+, Zn2+, Cd2+, Cu2+, Pb2+, Fe3+, and Al3+.
Typically, Ca2+ may be used. The alginate can be of any type,
including type G (L-guluronic acid) or type M (D-mannuronic acid),
or mixed M and G. For more information on alginate, see U.S. Pat.
No. 5,144,016.
[0043] In certain embodiments, a second polysaccharide, such as
dextran, can be physically trapped in the network formed by the
ionic cross-linking of the first polysaccharide. Dextran can be in
the form of covalently cross-linked beads, e.g., dextran that has
been previously cross-linked to itself with epichlorohydrin, or
Sephadex.TM. beads. Alternatively, the dextran can be in solution
form (e.g., uncross-linked), as described above. In addition,
dextran can be covalently linked to the cellulose, e.g. by linking
dextran to the cellulose with epichlorohydrin. Accordingly, dextran
may become cross-linked to itself.
[0044] Other embodiments of hemostatic compositions include
dextran-alginate spheres, such as ionically linked dextran-alginate
spheres, or covalently linked dextran-alginate spheres, or both
ionically and covalently linked dextran-alginate spheres. In
addition, cross-linked dextran spheres loaded with Ca2+ ions are
also included as hemostatic compositions of the present
invention.
[0045] Hemostatic compositions can include additional agents, such
as analgesics, steroids, antihistamines, anesthetics, bactericides,
disinfectants, fungicides, vasoconstrictors, hemostatics,
chemotherapeutic drugs, antibiotics, keratolytics, cauterizing
agents, antiviral drugs, epidermal growth factor, fibroblast growth
factors, transforming growth factors, glycoproteins, collagen,
fibrinogen, fibrin, humectants, preservatives, lymphokines,
cytokines, odor controlling materials, vitamins, and clotting
factors. For further information on these additional agents for
incorporation, refer to WO 00/27327.
[0046] Hemostatic compositions may be used in combination with
polymeric laminating materials and adhesives to provide both
mechanical support and flexibility to an article and to facilitate
adhesion to the wound. Additional information on such polymeric
laminating materials and adhesives for use in the present invention
can be found in, e.g., WO 00/27327.
[0047] Pharmaceutical Compositions
[0048] The present invention also contemplates pharmaceutical
compositions comprising certain hemostatic compositions of the
present invention, e.g., dextran-alginate linked spheres or
calcium-loaded cross-linked dextran spheres. Pharmaceutical
compositions may be formulated in conventional manners using one or
more physiologically acceptable carriers containing excipients and
auxiliaries. Proper formulation is dependent upon the route of
administration chosen.
[0049] A "pharmaceutically acceptable carrier" (also referred to
herein as an "excipient") is a pharmaceutically acceptable solvent,
suspending agent, or any other pharmacologically inert vehicle for
delivering one or more hemostatic compositions to a subject.
Pharmaceutically acceptable carriers can be liquid or solid, and
can be selected with the planned manner of administration in mind
so as to provide for the desired bulk, consistency, and other
pertinent transport and chemical properties, when combined with a
hemostatic composition. Other components may be present in a
pharmaceutical composition, if desired.
[0050] Pharmaceutical compositions of the present invention can be
administered by a number of methods depending upon the area to be
treated. Administration can be, for example, topical or parenteral.
Administration can be rapid (e.g., by injection) or can occur over
a period of time. For treating tissues in the central nervous
system, pharmaceutical compositions can be administered by
injection or infusion into the cerebrospinal fluid, preferably with
one or more agents capable of promoting penetration of the
pharmaceutical composition across the blood-brain barrier.
[0051] Formulations for topical administration include, for
example, sterile and non-sterile aqueous solutions, non-aqueous
solutions in common solvents such as alcohols, or solutions in
liquid or solid oil bases. Such solutions also can contain buffers,
diluents and other suitable additives. Pharmaceutical compositions
and formulations for topical administration can include patches,
ointments, lotions, creams, gels, drops, suppositories, sprays,
liquids, and powders. Conventional pharmaceutical carriers,
aqueous, powder or oily bases, thickeners and the like may be
necessary or desirable.
[0052] Compositions and formulations for parenteral administration
can include sterile aqueous solutions, which also can contain
buffers, diluents and other suitable additives (e.g., penetration
enhancers, carrier compounds and other pharmaceutically acceptable
carriers).
[0053] Pharmaceutical compositions of the present invention
include, but are not limited to, solutions, emulsions, aqueous
suspensions, and liposome-containing formulations. These
compositions can be generated from a variety of components that
include, for example, preformed liquids, self-emulsifying solids
and self-emulsifying semisolids. Emulsions are often biphasic
systems comprising of two immiscible liquid phases intimately mixed
and dispersed with each other; in general, emulsions are either of
the water-in-oil (w/o) or oil-in-water (o/w) variety. Emulsion
formulations have been widely used for oral delivery of
therapeutics due to their ease of formulation and efficacy of
solubilization, absorption, and bioavailability.
[0054] Pharmaceutical compositions of the invention further
encompass any pharmaceutically acceptable salts, esters, or salts
of such esters, or any other compound which, upon administration to
an animal including a human, is capable of providing (directly or
indirectly) the biologically active metabolite or residue thereof.
The term "pharmaceutically acceptable salts" refers to
physiologically and pharmaceutically acceptable salts of the
hemostatic compositions of the invention (i.e., salts that retain
the desired biological activity without imparting undesired
toxicological effects). Examples of pharmaceutically acceptable
salts include, but are not limited to, salts formed with cations
(e.g., sodium, potassium, calcium, or polyamines such as spermine);
acid addition salts formed with inorganic acids (e.g., hydrochloric
acid, hydrobromic acid, sulfuric acid, phosphoric acid, or nitric
acid); salts formed with organic acids (e.g., acetic acid, citric
acid, oxalic acid, palmitic acid, or fumaric acid); and salts
formed from elemental anions (e.g., chlorine, bromine, and
iodine).
[0055] Methods of Controlling Bleeding
[0056] In one aspect of the invention, a method for controlling
bleeding at an active bleeding wound site of an animal is provided.
The method includes applying a hemostatic composition to the active
bleeding wound site. Application of the hemostatic composition
typically includes contacting the hemostatic composition with the
wound or bleeding site surface. The hemostatic composition is
maintained in contact with the wound or bleeding site for a period
of time sufficient to control the bleeding, e.g., to clot the
blood, slow the rate of bleeding, or stop the bleeding. The
application may include the use of pressure, e.g., by using an
elastic bandage to maintain contact with the bleeding site.
Alternatively, an internal wound may be packed with a hemostatic
composition until hemostasis is achieved. In other embodiments, a
hemostatic composition is delivered to the wound site. For example,
a catheter or needle may be used to deliver a hemostatic
composition to an intravascular puncture site or to a biopsy site.
The catheter or the needle may be optionally coated with a
hemostatic composition of the present invention.
[0057] Usually a hemostatic composition can control bleeding, for
example, to a rate of less than 0.03 ml/min, in a period of from
about 2 to about 20 minutes. In certain embodiments, bleeding stops
immediately, or in less than about 5 minutes.
[0058] Typically a hemostatic compositions of the present invention
will be used to inhibit or completely stop bleeding of a
parenchymal organ, such as the liver, kidney, spleen, pancreas, or
lungs; or to control bleeding during surgery (e.g., abdominal,
vascular, gynecological, dental, tissue transplantation surgery,
etc.). For example, percutaneous needle biopsies are common
interventional medical procedures. Possible complications of needle
biopsies, however, include bleeding at the biopsy site. The amount
of bleeding is related to the needle size, tissue sample size,
location of the biopsy, and vascularization of the tissue.
Hemostatic compositions of the present invention can be used to
promote hemostasis at needle biopsy sites. Biopsy needles may
either be coated with hemostatic compositions of the present
invention, or may be used to deliver a hemostatic composition to
the biopsy site. For more information on biopsy tracts, see U.S.
Pat. No. 6,447,534.
[0059] Similarly, catheterization and interventional procedures,
such as angioplasty and stenting, generally are performed by
inserting a hollow needle through a patient's skin and muscle
tissue into the vascular system. A guide wire is then typically
passed through the needle lumen into a blood vessel. The needle is
removed and an introducer sheath is advanced over the guide wire
into the vessel, and a catheter is typically passed through the
lumen of the introducer sheath and advanced over the guide wire for
positioning. Upon completion of the medical procedure, the catheter
and introducer sheath are removed, often leaving a puncture site in
the vessel, with associated bleeding. Hemostatic compositions of
the present invention may be used to coat the exterior of
catheters, stents, introducer sheath, and guide wires, etc., or may
be delivered, e.g., via a catheter, to the puncture site in order
to promote hemostasis. For additional information, see U.S. Pat.
No. 6,391,048.
[0060] The amount of hemostatic composition to be used will vary
with the patient, the wound, and the composition employed. For
example, hemostatic compositions with varying water regains can be
assembled (e.g., stacked in descending order) for use in major
bleeding to attain hemostasis.
[0061] Methods for Making Hemostatic Compositions
[0062] In another aspect, the invention provides methods for making
hemostatic compositions. The hemostatic compositions of the present
invention can be made by incubating a linking agent with a
polysaccharide and an article containing cellulose to form a
hemostatic composition having the polysaccharide covalently linked
to the cellulose.
[0063] Any biologically compatible bifunctional or
heterobifunctional reagent may be used as the linking agent,
including reagents with halogens, epoxides, hydroxy succinimide
esters, aldehydes, activated thiols, or other moieties for reacting
free amines, hydroxides, hydroxyls, or sulfhydryls on the bandage
or on the polysaccharide. The bandage may be modified, e.g.,
derivatized, to incorporate reactive moieties such as amines or
sulfhydryls for reacting with a particular linking agent. The
polysaccharide may also be modified, e.g., derivatized, in a
similar manner, provided that the polysaccharide so derivatized
remains pharmaceutically suitable for animal, e.g., human use. The
linking agent may be epichlorohydrin, dichlorohydrin, diepoxyburan,
disepoxypropyl ether, or ethylene-glyco-bis-epoxypropylether. For
additional information, see Flodin, P., and Ingelman, B., "Process
for the Manufacture of Hydrophilic High Molecular Weight
Substances," British Patent No. 854,715; and Flodin, P., "Chapter
2: The Preparation of Dextran Gels," Dextran Gels and Their
Applications in Gel Filtration, Pharmacia, Uppsala Sweden, 1962,
pages 14-26.
[0064] The incubation step may occur in an aqueous alkaline
solution. Typically, the polysaccharide is from about 10% to about
80% wt/vol of the aqueous alkaline solution. The concentration of
the linking agent in the incubation step can range from about 2% to
about 20% wt/wt of the polysaccharide.
[0065] The temperature of the incubation step can range from about
40.degree. C. to about 70.degree. C. In certain embodiments, the
temperature is about 50.degree. C. The incubation step can occur
for about 1 to about 24 hours. The incubating step may also include
agitation of the reagents. In addition, the incubation step can be
in the presence of a stabilizing solution, e.g., a solution
designed to prevent or limit evaporation of water. The stabilizing
solution can include cellulose acetate butyrate. The method can
also include neutralizing the aqueous alkaline solution, e.g., with
an acid such as HCl at a concentration of from 1 to 5M.
[0066] The hemostatic composition can be washed with an aqueous
solution, e.g., distilled water, or an aqueous alcoholic wash,
e.g., 50/50 vol/vol EtOH/water. The hemostatic composition can be
serially washed in increasing amounts of an alcoholic wash, such as
25%, 50%, 75%, and 100% EtOH. The alcohol wash solution can contain
a humectant, e.g., glycerin, at a concentration of about 0.1 to
about 2.0%. The hemostatic composition can be dried, e.g., at about
50.degree. C. to about 80.degree. C. For example, the hemostatic
composition can be dried at 70.degree. C. After drying, the
covalently linked polysaccharide may have a molecular weight
exclusion limit of greater than 30,000 Daltons.
[0067] In certain embodiments of the method, the polysaccharide is
dextran. The dextran can be in the form of covalently cross-linked
beads. The molecular weight of the dextran can range from about
10,000 to about 2M, or from about 20,000 to about 100,000 Daltons.
Typically, dextran of MW 40,000 is used. The incubation step may be
occur in an aqueous alkaline solution having about 12 to about 75%
dextran wt/vol.
[0068] In another aspect, the invention provides a method of making
a hemostatic composition including incubating a polysaccharide and
a cation with an article containing cellulose in order to form a
hemostatic composition having the article containing cellulose in
association with an ionically cross-linked polysaccharide. The
polysaccharide may be further covalently linked to the cellulose.
The cation may be as described previously, including, for example,
Ca2+. The Ca2+ may be in the form of, or derived from, Ca2+-loaded
cross-linked, dextran beads. The polysaccharide may be sodium
alginate or derivatives of alginic acid, including salts of alginic
acid. Aqueous and alcoholic washes of the hemostatic composition
can be performed, as described previously.
[0069] In certain embodiments, the incubation step includes a
second polysaccharide. The second polysaccharide may be dextran,
e.g., dextran in the form of covalently cross-linked beads. The
second polysaccharide may be physically trapped, e.g., in the
three-dimensional network formed by the ionic cross-linking of the
first polysaccharide. The second polysaccharide may be further
covalently linked to the bandage, e.g., through a linking agent
such as epichlorohydrin.
[0070] In one embodiment, an article such as cellulose gauze is
immersed in a solution of a first polysaccharide (e.g., about 1 to
5% sodium alginate) and a second polysaccharide (e.g., 20% dextran,
avg. molecular weight 40,000). The first polysaccharide is
ionically cross-linked with a cation solution, e.g., Ca2+ from a
solution having about 0.5 to about 10% aqueous calcium chloride.
The Ca2+ concentration can be reduced with serial washes, e.g., to
reduce the Ca2+ concentration to about 0.5% Ca2+. The first or
second polysaccharide, or both, may then be covalently linked to
the cellulose and/or cross-linked using an aqueous alkaline
solution (e.g., 20% NaOH) of a linking agent, e.g., epichlorohydrin
(e.g., at about 3-6% of the weight of the second polysaccharide).
The resulting hemostatic composition may be dried as described
previously.
[0071] In another embodiment of the present invention, cross-linked
polysaccharide spheres can be mixed with a Ca2+-alcoholic wash
solution (e.g., 1% calcium chloride in neat alcohol). The
cross-linked polysaccharide spheres can be purchased, e.g., as
Sephadex.TM., or can be prepared from an aqueous alkaline
polysaccharide solution and a cross-linking agent (e.g., dextran
cross-linked with epichlorohydrin), as discussed previously. After
washing in the Ca2+-alcohol solution, the cross-linked
polysaccharide spheres have Ca2+ in their pores, e.g., are
Ca2+-loaded spheres, and can be used to coat an article, e.g.,
sprayed onto a bandage, that has been previously soaked or immersed
in a polysaccharide solution, e.g., sodium alginate, at a
concentration of about 0.5-5% polysaccharide. In certain
embodiments, Ca2+ from the cross-linked spheres exchanges with the
sodium from the sodium alginate solution, resulting in ionically
cross-linked calcium alginate, which physically traps cross-linked
polysaccharide spheres in the three-dimensional network of ionic
bonding. The hemostatic composition so formed may be dried as
discussed previously.
[0072] In another aspect, the invention provides a method for
manufacturing a composition, where the method includes the step of
mixing an aqueous phase alkaline polysaccharide solution with an
organic phase stabilizing agent solution to form a mixture having
polysaccharide spheres; incubating a cross-linking agent with the
mixture to cross-link the polysaccharide spheres; isolating the
cross-linked polysaccharide spheres; and coating an article
comprising a sodium alginate solution with the cross-linked
polysaccharide spheres. The method can include removing the organic
phase stabilizing agent from the mixture, e.g., prior to isolating
the cross-linked polysaccharide spheres. The method can also
include exposing the cross-linked polysaccharide spheres to a
solution comprising Ca2+ ions, e.g., washing the cross-linked
polysaccharide spheres in a Ca2+ solution. The polysaccharide may
be dextran, and the organic phase stabilizing agent solution may
include cellulose acetate butyrate.
[0073] In certain embodiments of the method, the cross-linked
polysaccharide spheres are between about 30 to about 500 .mu.m in
size. The mixing and incubating steps may occur at a temperature of
from about 40.degree. C. to about 70.degree. C. The coating step
can include spraying the article with the cross-linked
polysaccharide spheres. The invention also relates to hemostatic
compositions manufactured according to the above method.
[0074] In a further aspect, another method for manufacturing a
composition is provided. The method includes the steps of providing
an aqueous phase alkaline solution having dextran and sodium
alginate therein; preparing dextran-alginate spheres from the
aqueous phase alkaline solution; and incubating the
dextran-alginate spheres with a linking agent to form linked
dextran-alginate spheres. The dextran-alginate spheres can be
prepared by any method conventional in the art, including the use
of a mechanical droplet generator or an air knife.
[0075] The linking agent may covalently or ionically link and/or
cross-link the dextran-alginate spheres. Accordingly, the linking
agent may be epichlorohydrin or a cation as described previously,
e.g., a Ca2+-containing salt. The dextran-alginate spheres may be
first linked with a cation linking agent, and then linked with an
epichlorohydrin linking agent, or vice versa. The dextran-alginate
spheres may be linked simultaneously with a cation linking agent
and an epichlorohydrin linking agent. The method can further
including coating, e.g., spraying, an article with the linked
dextran-alginate spheres. The invention also is directed to
hemostatic compositions manufactured according to the method.
[0076] A number of embodiments of the invention have been
described. Nevertheless, it will be understood that various
modifications may be made without departing from the spirit and
scope of the invention. Accordingly, other embodiments are within
the scope of the following claims.
EXAMPLES
Example 1
Cotton-Dextran Compositions
[0077] Pharmaceutically acceptable cotton-based compositions were
incubated with dextran (40,000 MW) in an alkaline epichlorohydrin
solution (20% dextran in NaOH wt/vol; epichlorohydrin at about 3 to
6% wt/wt dextran). The solutions were allowed to react for about 1
to about 16 hours at a temperature range from about ambient room
temperature to about 60.degree. C. The resulting cross-linking
reactions were subsequently neutralized using a 1 to 5 Molar HCl
solution.
[0078] The cross-linked and linked dextran-cotton hemostatic
compositions were washed about 4 times with distilled water. The
products were further washed twice with a 50% distilled
water/alcohol solution, then with a 75% alcohol solution, and
lastly with a 100% alcohol solution, to remove excess
epichlorohydrin. A final alcohol wash solution contained about 0.1
to about 2% glycerin to keep the composition from becoming brittle.
The hemostatic composition was dried at about 70.degree. C.
overnight.
Example 2
Cotton-Alginate-Dextran Compositions
[0079] A stabilizing agent, such as cellulose acetate butyrate was
dissolved in ethylene dichloride at 3% wt/vol and heated to about
50.degree. C. while stirring at about 200 RPM in a 1-2 liter
cylindrical reaction vessel. Dextran (MW 40,000) was dissolved in
water at 15% wt/vol with 5N NaOH at 2% of the dextran weight. The
dextran solution was gradually added to the stabilizing mixture
with continued heating and stirring. When droplets of the desired
size were formed (30-500 .mu.m), a cross-linking agent such as
epichlorohydrin was added to the vessel at 20% of the dextran
weight. The reaction formed cross-linked gel spheres in 1-3 hours,
but was allowed to proceed up to 16 hours before termination.
Acetone was added and decanted twice to remove the stabilizer
(cellulose acetate butyrate). The spheres were then treated with
NaOH (equal parts of 2N NaOH and 95% ethyl alcohol) for about 15
mins., and neutralized with dilute acid (1N HCl) before filtration
and washing with water. The swollen spheres were shrunk with
alcohol treatment (25, 50, 75, 100% serial alcohol washes).
[0080] Dry calcium chloride was mixed with a second 100% alcohol
wash solution (1% calcium chloride in alcohol), which was used to
wash the cross-linked dextran particles. The alcohol was evaporated
off, trapping the calcium in the pores of the dextran particles.
The final products were dry cross-linked dextran-calcium ion
compositions.
[0081] Pharmaceutically acceptable cotton-based compositions were
immersed (dipped) into a sodium alginate liquid solution (0.5-4%).
After removal from the solution, the wet sodium alginate coated
cotton materials were sprayed or dusted with the cross-linked
dextran calcium compositions. Calcium exchanged with sodium,
resulting in cross-linked calcium alginate. The
cotton-dextran-alginate hemostatic compositions were then dried at
70.degree. C. overnight.
Example 3
Alternative Method to Prepare Cotton-Dextran-Alginate
Compositions
[0082] Cotton gauzes were immersed in solutions of 1 to 5% sodium
alginate and 20% dextran (40,000 MW). The mixtures were
cross-linked and linked with about 0.5% to about 10% aqueous
calcium chloride solution. The compositions were washed to reduce
Ca2+ concentration to about 0.5% Ca2+.
[0083] Dextran-alginate cross-linking solutions were prepared using
an aqueous alkaline epichlorohydrin solution, where the
concentration of the epichlorohydrin was from about 3 to about 6%
by weight of the dextran. The solutions included about 20% NaOH.
The resulting hemostatic compositions were allowed to dry at
ambient to about 60.degree. C. overnight.
Example 4
Cotton-Dextran Compositions
[0084] Dextran (MW 40,000) was dissolved in 1N NaOH at a range of
18-69% wt/vol of dextran in the alkaline solution. Epichlorohydrin
was added to a concentration of 20% of the dextran by weight at
room temperature. Pharmaceutically acceptable cotton (cellulose)
gauzes were added to the alkaline epichlorohydrin solutions. The
solutions were allowed to react with agitation at 25.degree. C. to
70.degree. C. The mixtures were heated until a cross-linked
dextran-cellulose gel formed on the gauze fibers, from 1-6 hours,
up to 24 hours. After neutralization with dilute HCl (1N),
successive washes removed excess reaction products and impurities:
four times in distilled water, then with increasing concentrations
of alcohol (25, 50, 75, 100%). A final alcohol wash of 100% EtOH
contained from 0.1 to 2.0% glycerin to keep the compositions
pliable. The resulting hemostatic compositions were dried at about
70.degree. C. overnight.
[0085] The water regain for the hemostatic compositions ranged from
2.5 ml/g to 35 ml/g. For information on water regain, see Flodin,
P., Dextran Gels and Their Applications in Gel Filtration,
Pharmacia, Uppsala Sweden, 1962, pages 31-32.
Example 5
Cellulose-Dextran Composition
[0086] 150 g of dextran (MW 40,000) was dissolved in 300 ml 1N
NaOH. 30 g epichlorohydrin was quickly mixed with the dextran
solution at room temperature. A pharmaceutically acceptable
cellulose gauze was dipped in the alkaline epichlorohydrin solution
to thoroughly coat the fibers with the reaction solution, then
placed in a flat-bottomed dish. The gauze was heated to 50.degree.
C. in a humidified atmosphere, with gentle rocking after 1 hour,
until a dextran-cellulose gel formed on the gauze fibers, typically
in 1-2 hours. Heating was continued until the desired end-point, up
to 24 hours. The gauze was neutralized, washed, treated with
glycerin, and dried as described previously. The water regain was
7.5 ml/g.
Example 6
Cellulose-Dextran Composition
[0087] Dextran (MW 40,000) was dissolved in 1N NaOH (34% dextran
wt/vol of the alkaline solution). Epichlorohydrin was added to a
concentration of 20% of the dextran by weight at room temperature.
A pharmaceutically acceptable cellulose fiber based composition,
16-ply 4.times.4 gauze, was dipped in the alkaline epichlorohydrin
solution to thoroughly coat the fibers with the reaction solution,
then placed in a flat bottomed dish. To prevent concentrating the
reaction solution by evaporation, a stabilizing solution of
cellulose acetate butyrate in ethylene dichloride (3% wt/vol),
which is immiscible in water, was used to cover the gauze. The
gauze was heated at 50.degree. C., with gentle rocking after 1 hr.,
until a cross-linked and linked dextran-cellulose gel formed on the
gauze fibers, typically in about 2 to 3 hours. Heating was
continued until the desired endpoint, up to 24 hours. Acetone was
added and decanted twice to remove the stabilizer. The gauze was
neutralized with dilute HCl and washed as described previously in
aqueous alcohol and alcohol solutions and dried at about 70.degree.
C. overnight. The water regain was 15 ml/g.
Example 7
Cellulose Dextran Composition
[0088] Dextran (MW 40,000) was dissolved in 1N NaOH and reacted
with gauze in the presence of epichlorohydrin, followed by
neutralization, washing, and drying as described above.
Cellulose-dextran compositions were made by varying the volume of
dextran in the solvent from 12 to 75 wt/vol to produce hemostatic
composition with water regains ranging from 5 ml/g to 35 ml/g. The
compositions can be assembled for use in major bleeding by stacking
them in descending order (e.g., 35 ml/g to 5 ml/g) to attain
hemostasis.
Example 8
Dextran-Calcium Spheres
[0089] A non-ionic polymer substance was dissolved in a suitable
solvent with an alkaline solution added as a cross-linking
catalyst. A stabilizer was dissolved in a solvent that was
immiscible with the polymer solvent and placed in a cylindrical
vessel. The stabilizer solution formed the continuous phase and was
heated with regular stirring. When the polymer solution was added
to the stabilizer solution, a biphasic system formed in which the
polymer droplets became the dispersed phase. A bifunctional
cross-linking agent was added to the system which caused
co-polymerization (cross-linking) of the polymer to form gel
spheres.
[0090] After purification and drying, the water regain of the
spheres was determined in order to classify the molecular sieving
capability of the cross-linked polymer. Water regain can be
determined by methods well known in the art, and generally involves
hydrating 1 gram of dry composition, and determining the amount of
water absorbed by the 1 g of the dry composition. Generally,
greater swelling capacity relates to larger pores (e.g., less
cross-linking) and a higher molecular weight exclusion.
[0091] More specifically, a stabilizing agent, such as cellulose
acetate butyrate was dissolved in ethylene dichloride at 3% wt/vol
and heated to about 50.degree. C. while stirring at about 200 RPM
in a 1-2 liter cylindrical reaction vessel. Dextran (MW 40,000) was
dissolved in water at 15% wt/vol with 5N NaOH at 2% of the dextran
weight. The dextran solution was gradually added to the stabilizing
mixture with continued heating and stirring. When droplets of the
desired size were formed (30-500 .mu.m), a cross-linking agent such
as epichlorohydrin was added to the vessel at 20% of the dextran
weight. The reaction formed cross-linked gel spheres in 1-3 hours,
but was allowed to proceed up to 16 hours before termination.
Acetone was added and decanted twice to remove the stabilizer
(cellulose acetate butyrate). The spheres were then treated with
NaOH (equal parts of 2N NaOH and 95% ethyl alcohol) for about 15
mins., and neutralized with dilute acid (1N HCl) before filtration
and washing with water. The swollen spheres were shrunk with
alcohol treatment (25, 50, 75, 100% serial alcohol washes) as
described previously. A final alcohol wash contained calcium
chloride (0.04-1%). The alcohol was evaporated by drying at about
70.degree. C., thus trapping the calcium in the pores and on the
surface of the dextran spheres. The final product was a dry
cross-linked dextran-calcium ion composition. The water regain was
20 ml/g.
Example 9
Cotton-Alginate-Dextran Compositions
[0092] A pharmaceutically acceptable cotton (cellulose) gauze was
immersed in or sprayed with a sodium alginate solution (0.5-4%).
The wet sodium alginate coated gauzes were sprayed or dusted with
dextran-calcium spheres, prepared as described previously. Calcium
exchanged with sodium in the alginate, resulting in cross-linked
calcium alginate. Ionic bonds formed between cellulose, alginate,
and dextran, thereby chemically incorporating the spheres into the
gauze to form the hemostatic compositions.
Example 10
Dextran-Alginate Spheres
[0093] 43 g dextran (MW 40,000) was dissolved in 50 ml 2N NaOH and
mixed with a 50 ml solution of 2% sodium alginate (43% dextran and
1% alginate in 100 ml of 1N NaOH). 50-200 .mu.m dextran-alginate
droplets were linked and cross-linked in a solution of 1.7% calcium
chloride. The spheres were further linked and cross-linked by
adding 7.2 ml epichlorohydrin to 100 ml of the calcium chloride
solution at 45.degree. C. with agitation. The reaction continued
for up to 16 hours until the desired amount of linking and
cross-linking occurred. The spheres were neutralized with dilute
HCl, washed in increasing concentrations of alcohol (25, 50, 75,
100%) and dried at about 70.degree. C. overnight. The water regain
was 10 ml/g.
Example 11
Cotton-Dextran-Alginate Sphere Composition
[0094] Cross-linked and linked dextran-alginate spheres, prepared
as described previously, were washed in a final calcium chloride
alcohol wash (0.04-1% calcium chloride). The alcohol was evaporated
to trap the calcium in the pores of the spheres. The spheres were
then chemically incorporated into gauze containing sodium alginate,
as described previously.
Example 12
Compositions having Dextrans of Different Molecular Weights
[0095] 100 g of dextran (MW 2,000,000) dissolved in 2000 ml 0.5N
NaOH was mixed with 30 g of dextran spheres, prepared as described
previously. The cross-linked spheres and dextran solutions were
mixed, and 100 g epichlorohydrin added. A pharmaceutically
acceptable cellulose gauze was placed in a flat bottomed dish and
the alkaline epichlorohydrin/dextran sphere/dextran solution was
slowly poured over the gauze to thoroughly coat the fibers with the
dextran spheres and dextran solution The coated gauze was heated at
50.degree. C. with gentle rocking, until a gel formed on the gauze
fibers, typically in 1-2 hours. The coated gauze was removed from
the solution and placed in a flat bottom dish and heating was
continued until the desired end-point, approximately 8-12 hours.
The gauze was neutralized, washed, and dried, as described
previously.
Example 13
Tests of Cotton-Alginate-Dextran Compositions and Calcium-Dextran
Spheres
[0096] Two different alginates (G, M) were applied in 0.5% and/or
1% concentrations to 12-ply 2.times.2 cotton gauze squares, Type
VII, in either 2 or 4 mL amounts. 0.5 g of calcium-dextran spheres
(containing 3, 6, 12, or 24 mM calcium) were ionically bonded to
the sodium alginate on the gauze. Freshly drawn human blood was
added to each test sample, with 0.5 mls added to each
calcium-dextran-alginate gauze and 0.1 ml added to the
calcium-dextran spheres alone. Gauze immersed only in sodium
alginate was used as a control.
[0097] The blood added to the calcium-dextran spheres clotted upon
contact. All of the calcium-dextran-alginate gauzes clotted the
blood in less than 5 minutes, whereas alginate only gauzes produced
weakly clotted blood in>5 mins. See Table 1 below.
TABLE-US-00001 TABLE 1 Clotting Times for Hemostatic Compositions
gauze + 4 gauze + 4 gauze + 2 gauze + 4 ml .5% ml 1% ml 1% ml 1%
alginate G alginate G alginate G alginate M 0 mM Ca++ >5 min.
>5 min. >5 min. >5 min. control 3 mM Ca++ instantly not
tested <5 min. not tested not tested dextran spheres 6 mM Ca++
instantly <5 min. <5 min. <5 min. <5 min. dextran
spheres 12 mM instantly <5 min. <5 min. <5 min. <5 min.
Ca++ dextran spheres 24 mM instantly <5 min. <5 min. <5
min. <5 min. Ca++ dextran spheres
* * * * *